Validation of the VM7Luc Test Method
In their responses to a 2012 ICCVAM report, U.S. federal agencies agreed with ICCVAM recommendations on the use of human cell-based test methods to screen for substances with the potential to interact with the estrogen receptor (ER).
ICCVAM evaluated the usefulness and limitations of the VM7Luc agonist and antagonist tests and made recommendations on how they could be used to identify substances that induce or inhibit human ER activity in vitro. The U.S. Environmental Protection Agency (EPA) responded that they regard these tests as an alternative to the ER transactivation (TA) test method used in their Endocrine Disruptor Screening Program. Several other federal agencies stated that they would communicate the ICCVAM recommendations to various stakeholders and encourage appropriate use of the recommended methods.
- Test Method Evaluation Report (2012) with ICCVAM recommendations
Change in Assay Designation (June 2016)
In 2016, NICEATM received new information regarding the LUMI-CELL® test. DNA testing has now shown that the original cell line used to develop the test was not the BG1 human ovarian carcinoma cell line. It was a variant of the MCF7 human breast cancer cell line. The cell line used in the test, originally referred to as BG1Luc4E2 (Rogers and Denison 2000), will now be designated as VM7Luc4E2 ("V" = variant; "M7" = MCF7 cells). Likewise, the test will now be generically referred to as the VM7Luc ER TA test. While this changes the origin of the cell line upon which the test is based, it does not affect published validation studies nor the use and application of this test for screening of estrogenic/anti-estrogenic chemicals.
This correction should accompany any information currently referencing the LUMI-CELL test (e.g., webpages, databases, documents, manuscripts).
- Supporting Documentation from Michael Denison, Department of Environmental Toxicology, University of California, Davis
Summary of Test Method Evaluation and Acceptance
ICCVAM developed the recommendations on the VM7Luc tests after considering comments from:
- A 2011 international independent peer review panel
- The public
- The Scientific Advisory Committee on Alternative Toxicological Methods
ICCVAM concluded that the accuracy of the VM7Luc agonist test was at least equivalent to the only ER TA test method in a U.S. regulatory test guideline: the EPA's OPPTS 890.1300: Estrogen Receptor Transcriptional Activation (Human Cell Line (HeLa-9903)). The VM7Luc tests offer several advantages over the existing ER TA method, including:
- Validation for use over a wider concentration range of test substances
- Potential to detect a wider range of ER-active substances
- Ability to identify both substances that induce and inhibit the estrogen receptor
- Availability of the cell line used for the test from more than one source
The ICCVAM recommendations on the VM7Luc tests formed the basis for Test Guideline 455 issued in 2012 and updated most recently in 2016 by the Organization for Economic Co-operation and Development (OECD). Adoption of Test Guideline 455 means that the VM7Luc tests can be used in the 35 OECD member countries to identify substances that induce or inhibit human ER activity in vitro.
Background on the VM7Luc Validation Study
The VM7Luc test was developed by Xenobiotic Detection Systems, Inc., under the name LUMI-CELL with support from NIEHS Small Business Innovation Research grant ES10533-03. The test method measures the extent to which a substance induces or inhibits TA activity via ER-mediated pathways in VM7Luc4E2 cells.
The VM7Luc4E2 cell line was derived from a variant of the MCF7 human breast cancer cell line that endogenously expresses both human ER forms, ERα and ERβ. The variant cell line contains four copies of a synthetic oligonucleotide containing the estrogen response element upstream of the mouse mammary tumor viral promoter. The firefly luciferase gene is used as a reporter, causing the VM7Luc4E2 cells to emit measurable light in response to estrogen and estrogen-like substances.
NICEATM coordinated an international interlaboratory validation study to evaluate the VM7Luc test method. It was the first validation study sponsored jointly by:
- The European Centre for the Validation of Alternative Methods (now known as the European Union Reference Laboratory for Alternatives to Animal Testing)
- The Japanese Center for the Validation of Alternative Methods
An independent international peer review panel met in 2011 to review the study results. They agreed with ICCVAM draft test method recommendations that an in vitro test method may be used as an initial screen to identify substances with the potential to enhance or inhibit activation of the estrogen receptor.
VM7Luc Test Method in the Tox21 Program
NICEATM nominated the VM7Luc agonist and antagonist tests for evaluation and adoption by the Tox21 program. The assays have now been translated to a high-throughput format using 1536-well plates by the National Center for Advancing Translational Sciences and have been used to screen all compounds in the Tox21 10K chemical library.
Transmittal of Recommendations to Federal Agencies
- National Institute of Environmental Health Sciences (NIEHS)
- Agency for Toxic Substances and Disease Registry (ATSDR)
- Consumer Product Safety Commission (CPSC)
- Environmental Protection Agency (EPA)
- Food and Drug Administration (FDA)
- National Cancer Institute (NCI)
- Letter from Dr. Birnbaum to Harold Varmus, M.D., Director, NCI
- Response from Dr. Varmus (received February 7, 2012)
- National Institutes of Health (NIH)
- Letter from Dr. Birnbaum to Francis S. Collins, M.D., Ph.D., Director, NIH
- Response from Dr. Collins (received June 26, 2012)
- National Institute for Occupational Safety and Health (NIOSH)
- Letter from Dr. Birnbaum to John Howard, M.D., Director, NIOSH
- Response from Dr. Howard (received June 26, 2012)
- National Library of Medicine (NLM)
- Occupational Safety and Health Administration (OSHA)
- U.S. Department of Agriculture
- U.S. Department of Defense
- U.S. Department of Energy
- U.S. Department of the Interior
- U.S. Department of Transportation
Other Materials Relevant to the Validation Study
- Test Method Nomination: Submission of XDS's LUMI-CELL ER High-Throughput System for Screening Estrogen-Like Chemicals for Review by ICCVAM (2005)
- Draft Evaluation: Nomination of the LUMI-CELL ER High-Throughput System for Screening Estrogen-Like Chemicals for Validation Studies (2004)
- Protocols for the International Validation Study